The goal of this proposal is to develop a universal method for rapidly deriving effective inhibitors for any gene of interest. Accomplishing this goal requires two steps: 1) selecting an accessible site on the mRNA, and 2) designing an effective molecule that will recognize that site and bind to it, or inactivate it by some other means. Currently available antisense technology and accessible target site selection is still in its infancy and unreliable. Therefore we propose a novel and general approach for rapidly developing an effective technology for mapping accessible target sites in any RNA as well as engineering effective inhibitors to those mapped accessible sites. We accomplish this by using random libraries of antisense sequences that, functioning inside the cell where the target is in its natural conformation, automatically identify the most accessible sites in any desired target RNA. By employing those identified accessible sites, we will engineer our proprietary, topology-based antisense design, RNA Padlocks, for targeted inhibition of mRNA of any desired gene. PROPOSED COMMERCIAL APPLICATIONS: Antisense technologies are currently hindered by the poor ability of oligonucleotides to bind to most sites on target mRNAs, and the difficulty of identifying those sites to which they can bind. By overcoming this hurdle, we will help realize the great possibilities of rational drug design and drug target validation that antisense technology originally promised.